Low-dose decitabine enhances the efficacy of viral cancer vaccines for immunotherapy.

Cancer immunotherapy requires a specific antitumor CD8+ T cell-driven immune response; however, upon genetic and epigenetic alterations of the antigen processing and presenting components, cancer cells escape the CD8+ T cell recognition. As a result, poorly immunogenic tumors are refractory to conventional immunotherapy. In this context, the use of viral cancer vaccines in combination with hypomethylating agents represents a promising strategy to prevent cancer from escaping immune system recognition. In this study, we evaluated the sensitivity of melanoma (B16-expressing ovalbumin) and metastatic triple-negative breast cancer (4T1) cell lines to FDA-approved low-dose decitabine in combination with PeptiCRAd, an adenoviral anticancer vaccine. The two models showed different sensitivity to decitabine in vitro and in vivo when combined with PeptiCRAd. In particular, mice bearing syngeneic 4T1 cancer showed higher tumor growth control when receiving the combinatorial treatment compared to single controls in association with a higher expression of MHC class I on cancer cells and reduction in Tregs within the tumor microenvironment. Furthermore, remodeling of the CD8+ T cell infiltration and cytotoxic activity toward cancer cells confirmed the effect of decitabine in enhancing anticancer vaccines in immunotherapy regimens.

Exploring the DNA2-PNA heterotriplex formation in targeting the Bcl-2 gene promoter: A structural insight by physico-chemical and microsecond-scale MD investigation.

Peptide Nucleic Acids (PNAs) represent a promising tool for gene modulation in anticancer treatment. The uncharged peptidyl backbone and the resistance to chemical and enzymatic degradation make PNAs highly advantageous to form stable hybrid complexes with complementary DNA and RNA strands, providing higher stability than the corresponding natural analogues. Our and other groups' research has successfully shown that tailored PNA sequences can effectively downregulate the expression of human oncogenes using antigene, antisense, or anti-miRNA approaches. Specifically, we identified a seven bases-long PNA sequence, complementary to the longer loop of the main G-quadruplex structure formed by the bcl2midG4 promoter sequence, capable of downregulating the expression of the antiapoptotic Bcl-2 protein and enhancing the anticancer activity of an oncolytic adenovirus. Here, we extended the length of the PNA probe with the aim of including the double-stranded Bcl-2 promoter among the targets of the PNA probe. Our investigation primarily focused on the structural aspects of the resulting DNA2-PNA heterotriplex that were determined by employing conventional and accelerated microsecond-scale molecular dynamics simulations and chemical-physical analysis. Additionally, we conducted preliminary biological experiments using cytotoxicity assays on human A549 and MDA-MB-436 adenocarcinoma cell lines, employing the oncolytic adenovirus delivery strategy.

Microbiome Profiling in Bladder Cancer Patients Using the First-morning Urine Sample.

Background: Several studies support the interplay between the urinary microbiome (ie, urobiome) and bladder cancer (BCa). Specific urinary bacteria may be responsible for chronic inflammation, which in turn promotes carcinogenesis. Different signatures of urobiome in BCa patients were identified depending on tumor type, geographical area, age, and sex. Objective: We explored the urobiome in BCa patients undergoing transurethral resection of bladder tumor (TURBT), to identify possible predictive biomarkers of cancer. Design setting and participants: The urobiome analysis was conducted in 48 patients (13 females) undergoing TURBT, of whom 30 with BCa (five females) and 18 with benign bladder tumor, analyzing bacterial 16S rRNA by next-generation sequencing in first-morning (FM) urine samples. Forty-three cancer-free individuals and 17 prostate cancer patients were used as controls. Outcome measurements and statistical analysis: First, we identified the better urine collection procedure to perform the urobiome analysis, comparing bacterial composition between catheterized (CAT) and FM urine samples in TURBT patients. Successively, we observed a specific urobiome in BCa patients rather than controls. A combined pipeline including the DESeq2 and linear discriminant analysis effect size tests was used to identify differential urinary taxa, strictly associated with BCa patients. Results and limitations: The bacterial composition of CAT and FM urine samples was comparable, so the latter was used for the following analyses. An increased abundance of Porphyromonas and Porphyromonas somerae was found in BCa patients compared with controls. This signature seems to be more related (p <0.05) to male BCa patients over 50 yr old. Owing to the low biomass of urinary microbiota, several samples were excluded from the study, reducing the number of BCa patients considered. Conclusions: FM urine samples represent a manageable specimen for a urobiome analysis; P. somerae is a specific biomarker of BCa risk. Patient summary: Our study showed an increased abundance of Porphyromonas and Porphyromonas somerae in male bladder cancer (BCa) patients, supporting the use of a first-morning urine sample, a less invasive and low-cost collection method, for the urobiome analysis of patients at risk of BCa.

Bifidobacterium affects antitumor efficacy of oncolytic adenovirus in a mouse model of melanoma.

Gut microbiota plays a key role in modulating responses to cancer immunotherapy in melanoma patients. Oncolytic viruses (OVs) represent emerging tools in cancer therapy, inducing a potent immunogenic cancer cell death (ICD) and recruiting immune cells in tumors, poorly infiltrated by T cells. We investigated whether the antitumoral activity of oncolytic adenovirus Ad5D24-CpG (Ad-CpG) was gut microbiota-mediated in a syngeneic mouse model of melanoma and observed that ICD was weakened by vancomycin-mediated perturbation of gut microbiota. Ad-CpG efficacy was increased by oral supplementation with Bifidobacterium, reducing melanoma progression and tumor-infiltrating regulatory T cells. Fecal microbiota was enriched in bacterial species belonging to the Firmicutes phylum in mice treated with both Bifidobacterium and Ad-CpG; furthermore, our data suggest that molecular mimicry between melanoma and Bifidobacterium-derived epitopes may favor activation of cross-reactive T cells and constitutes one of the mechanisms by which gut microbiota modulates OVs response.

Targeted inhibition of the methyltransferase SETD8 synergizes with the Wee1 inhibitor adavosertib in restraining glioblastoma growth.

Despite intense research efforts, glioblastoma remains an incurable brain tumor with a dismal median survival time of 15 months. Thus, identifying new therapeutic targets is an urgent need. Here, we show that the lysine methyltransferase SETD8 is overexpressed in 50% of high-grade gliomas. The small molecule SETD8 inhibitor UNC0379, as well as siRNA-mediated inhibition of SETD8, blocked glioblastoma cell proliferation, by inducing DNA damage and activating cell cycle checkpoints. Specifically, in p53-proficient glioblastoma cells, SETD8 inhibition and DNA damage induced p21 accumulation and G1/S arrest whereas, in p53-deficient glioblastoma cells, DNA damage induced by SETD8 inhibition resulted in G2/M arrest mediated by Chk1 activation. Checkpoint abrogation, by the Wee1 kinase inhibitor adavosertib, induced glioblastoma cell lines and primary cells, DNA-damaged by UNC0379, to progress to mitosis where they died by mitotic catastrophe. Finally, UNC0379 and adavosertib synergized in restraining glioblastoma growth in a murine xenograft model, providing a strong rationale to further explore this novel pharmacological approach for adjuvant glioblastoma treatment.

Systems Biology Approaches for the Improvement of Oncolytic Virus-Based Immunotherapies.

Oncolytic virus (OV)-based immunotherapy is mainly dependent on establishing an efficient cell-mediated antitumor immunity. OV-mediated antitumor immunity elicits a renewed antitumor reactivity, stimulating a T-cell response against tumor-associated antigens (TAAs) and recruiting natural killer cells within the tumor microenvironment (TME). Despite the fact that OVs are unspecific cancer vaccine platforms, to further enhance antitumor immunity, it is crucial to identify the potentially immunogenic T-cell restricted TAAs, the main key orchestrators in evoking a specific and durable cytotoxic T-cell response. Today, innovative approaches derived from systems biology are exploited to improve target discovery in several types of cancer and to identify the MHC-I and II restricted peptide repertoire recognized by T-cells. Using specific computation pipelines, it is possible to select the best tumor peptide candidates that can be efficiently vectorized and delivered by numerous OV-based platforms, in order to reinforce anticancer immune responses. Beyond the identification of TAAs, system biology can also support the engineering of OVs with improved oncotropism to reduce toxicity and maintain a sufficient portion of the wild-type virus virulence. Finally, these technologies can also pave the way towards a more rational design of armed OVs where a transgene of interest can be delivered to TME to develop an intratumoral gene therapy to enhance specific immune stimuli.

8-Hydroxy-2-Deoxyguanosine and 8-Iso-Prostaglandin F2α: Putative Biomarkers to assess Oxidative Stress Damage Following Robot-Assisted Radical Prostatectomy (RARP).

Objective: Prostate cancer (PCa) is the most common type of cancer. Biomarkers help researchers to understand the mechanisms of disease and refine diagnostic panels. We measured urinary 8-hydroxy-2-deoxyguanosine (8-OHdG) and 8-iso-prostaglandin F2α (8-IsoF2α) to assess oxidative stress damage in PCa patients undergoing robot-assisted radical prostatectomy (RARP). Methods: Forty PCa patients were enrolled in the study. Urine was collected before (T0) and 3 months after the RARP procedure (T1). 8-OHdG and 8-IsoF2α were measured through liquid chromatography-tandem mass spectrometry. Sex- and age-matched healthy subjects served as controls (CTRL). Results: At T0, patients exhibited significantly higher levels of 8-OHdG than CTRL (p = 0.026). At T1, 23/40 patients who completed the 3-month follow-up showed levels of 8-OHdG that were significantly lower than at T0 (p = 0.042), and comparable to those of the CTRL subjects (p = 0.683). At T0, 8-Iso-PGF2α levels were significantly higher in PCa patients than in CTRL subjects (p = 0.0002). At T1, 8-Iso-PGF2α levels were significantly lower than at T0 (p < 0.001) and were comparable to those of CTRL patients (p = 0.087). Conclusions: A liquid chromatography-tandem mass spectrometry method reveals enhanced OHdG and 8-Iso-PGF2α in the urine of PCa patients. RARP normalizes such indices of oxidative stress. Large-sized sample studies and long-term follow-ups are now needed to validate these urinary biomarkers for use in the early prevention and successful treatment of PCa.

Oncolytic Adenoviral Vector-Mediated Expression of an Anti-PD-L1-scFv Improves Anti-Tumoral Efficacy in a Melanoma Mouse Model.

Oncolytic virotherapy is an emerging therapeutic approach based on replication-competent viruses able to selectively infect and destroy cancer cells, inducing the release of tumor-associated antigens and thereby recruiting immune cells with a subsequent increase in antitumoral immune response. To increase the anticancer activity, we engineered a specific oncolytic adenovirus expressing a single-chain variable fragment of an antibody against PD-L1 to combine blockage of PD-1/PD-L1 interaction with the antitumoral activity of Onc.Ad5. To assess its efficacy, we infected B16.OVA cells, a murine model of melanoma, with Ad5Δ24 -anti-PD-L1-scFv and then co-cultured them with C57BL/6J naïve splenocytes. We observed that the combinatorial treatments were significantly more effective in inducing cancer cell death. Furthermore, we assessed the efficacy of intratumoral administrations of Ad5Δ24-anti-PD-L1-scFv in C57BL/6J mice engrafted with B16.OVA and compared this treatment to that of the parental Ad5Δ24 or placebo. Treatment with the scFv-expressing Onc.Ad induced a marked reduction of tumor growth concerning the parental Onc.Ad. Additionally, the evaluation of the lymphocytic population infiltrating the treated tumor reveals a favorable immune profile with an enhancement of the CD8+ population. These data suggest that Onc.Ad-mediated expression of immune checkpoint inhibitors increases oncolytic virotherapy efficacy and could be an effective and promising tool for cancer treatments, opening a new way into cancer therapy.

Oncolytic Adenoviruses for Cancer Therapy.

Many immuno-therapeutic strategies are currently being developed to fight cancer. In this scenario, oncolytic adenoviruses (Onc.Ads) have an interesting role for their peculiar tumor selectivity, safety, and transgene-delivery capability. The major strength of the Onc.Ads is the extraordinary immunogenicity that leads to a strong T-cell response, which, together with the possibility of the delivery of a therapeutic transgene, could be more effective than current strategies. In this review, we travel in the adenovirus (Ads) and Onc.Ads world, focusing on a variety of strategies that can enhance Onc.Ads antitumoral efficacy, passing through tumor microenvironment modulation. Onc.Ads-based therapeutic strategies constitute additional weapons in the fight against cancer and appear to potentiate conventional and immune checkpoint inhibitors (ICIs)-based therapies leading to a promising scenario.